Abstract Ovarian cancer (OC) is deadly and incurable for patients that recur after primary cancer is treated with surgery and platinum-based chemotherapy. Unfortunately, OC has a high rate of disease recurrence (70-80%) and most recurrent tumors are chemoresistant. Currently, therapeutic options are quite limited for these patients, and thus unlike with other cancers, the mortality in OC has not improved in decades. This dreadful situation highlights a drastic need for new and improved therapeutics to eliminate and treat recurrent and chemoresistant disease. In this context our studies identified increased Hedgehog (Hh)/GLI signaling in chemoresistant OC cells and tumors, and GLI1 expression significantly correlated with upregulated FA-BRCA and homologous recombination (HR) genes, cancer stem cell signaling and poor prognosis of OC patients. Our preliminary data demonstrate that GLI1 regulates expression of DNA repair genes BRCA1, FANCD2 and RAD51 in OC cells and tumors and its inhibition induces a state known as ?BRCAness?, which is characterized by inefficient repair of DNA double strand breaks (DSB). Additionally, Hh/GLI1 inhibition or downregulation decreased cell survival and ability to form tumor spheroids (stem cell characteristic), and enabled synthetic lethality with PARP inhibitors. In this proposal we aim to elucidate the molecular mechanisms by which Hh/GLI1 regulates FA-BRCA and HR genes that promote oncogenic replication (Aim 1), and determine the efficacy of Hh/GLI1 inhibitors in both BRCA-proficient and deficient OC cells in cell line and ovarian cancer orthotopic mouse models (Aim 2). Towards the goal of identifying effective therapeutic combinations that work synergistically and induce synthetic lethality, we will evaluate effective therapeutic combinations involving Hh/GLI1 and PARP inhibitors. The most efficient drug combinations will be examined in a mouse orthotopic and patient derived xenograft models of OC to evaluate their efficacy in combatting acquired chemoresistance and OC disease progression (Aim 3). Chemotherapeutic drugs examined in this proposal are FDA approved or currently undergoing clinical and preclinical evaluation (ex. vismodegib, sonidegib, GANT61, olaparib, rucaparib, niraparib); thus, the effective drug combinations discovered can quickly be transitioned into clinical trials and developed into treatments for OC patients.